The purpose of the proposed research is to isolate and analyze transcribed, repeated sequences (i.e., 5S DNA and ribosomal DNA) from Chinese hamster cells. Information regarding the structure and organization of these DNAs will be utilized to further our understanding of how these genes replicate. The 5S DNA appears heterogeneous with respect to base composition of the non-transcribed spacer. One population of 5S DNA bands in a light position in actinomycin D-CsCl gradients and will be isolated by isopycnic centrifugation. The repeat size of the DNA will be determined by electron microscopy and by restriction enzyme digestion. We will determine when this 5S DNA replicates within the S phase and attempt to isolate replicating molecules of 5S DNA from synchronized cells. Electron microscopy of such molecules should reveal whether replication initiation sites are randomly distributed or whether they occur at intervals equal to a repeat length or at multiples thereof. The 5S DNA from the main band will also be isolated, but in single stranded form, and the two populations will be compared by heteroduplex mapping. The rDNA will be isolated by two methods and studied with respect to repeat length. Heterogeneity already described will be further analyzed.